This work aims at providing a more detailed understanding of the function and mechanism of the enzyme DNA ligase from Escherichia coli. This objective will be approached by synthesizing several NAD ion analogues in which the oxygen atom attached to the 5'-carbon atom of the adenylic acid moiety of NAD ion has been substituted by less electronegative groups ranging from sulfur through nitrogen to the methylene group. In addition, procedures for the large-scale and rapid purification on DNase and RNase free bacteriophage T4-induced RNA ligase will be developed. Several reactions of the purified enzyme will be studied including the transfer of the non-adenylyl portion of molecules of the general structure Ado-5'-PP-X to nucleic acids. The RNA ligase catalyzed joining of single stranded oligodeoxyribonucleotides will be optimized. BIBLIOGRAPHIC REFERENCE: Bedows, E., Wachsman, J.T., and Gumport, R.I., Biochemistry, 16, 2231 (1977) (reprints attached) "L Cell DNA Ligase Joins RNA to DNA on a DNA Template."